Freshly-frozen ‘WakeField’ red raspberry (RR) fruits were generously donated by Enfield Farms (Lynden, WA). The fractionation and extraction of RR polyphenols from whole fruit, seed, and pulp were conducted according to Gourineni et al. [15 (link)]. Briefly, the physical separation of pulp and seed fractions was performed by using a fine sieve. The RR polyphenols from each fraction (i.e., whole fruits, pulp, or seed) were extracted with acidified methanol (0.5% acetic acid). After removing sugar and fibers by macroporous ion-exchange resin column (Ambelite FPX66, Dow Chemical), each polyphenolic fraction was dried, lyophilized, and stored at −20 °C. Approximately, 25 g whole RR polyphenols were obtained from 6 kg of freshly frozen fruit (~410 mg/100g RR). Upon fractionation of 6 kg frozen RR, 15 g of pulp polyphenol and 5 g of seed polyphenols were obtained. The composition of RR polyphenolic compounds was analyzed on an Agilent 1200 HPLC (Agilent Zorbax SB-C18 column) equipped with a binary pump, injector, diode array detector, and fluorescent detector (Agilent Technologies) as we have described previously [16 ]. Total phenolic content was determined by using Folin-Ciocalteu reagent with gallic acid as a standard (GAE, gallic acid equivalent). The composition of RR polyphenols is summarized in Table 1 .
Fluorescent detector
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The Fluorescent detector is a sensitive analytical instrument used to detect and measure the presence of fluorescent compounds in a sample. It operates by exciting the sample with a specific wavelength of light, causing the molecules to emit light at a longer wavelength, which is then detected and quantified.
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2 protocols using fluorescent detector
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Extraction and Characterization of Raspberry Polyphenols
2
Tocopherol Analysis in Olive Oil
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Sample preparation for the analysis of tocopherols in extra virgin olive oil was performed prior to injection to the LiChrospher Si60 column (inner diameter 25 cm, length 4.6 mm and particle size 5 µm) (Merck, Darmstadt, Germany), fitted in an Agilent 1200 high performance liquid chromatography (HPLC) system equipped with a fluorescent detector (Agilent Technologies, Santa Clara, CA, USA). The sample preparation method is herein: 0.15 g of olive oil was diluted in 10 mL of n-hexane followed by centrifugation with a rotor speed of 25,000× g for 10 min in a laboratory cold centrifuge (HERMLE Labortechnik, Wehingen, Germany). The supernatant was transferred to a chromatographic vial and 20 µL of the sample was injected into the HPLC system. Chromatographic separation was performed by the isocratic mixture of isopropanol:hexane 0.5:99.5 (v/v) mobile phase, operated with a 0.7 mL·min−1 flow rate. The fluorescence detector was set to excitation and emission wavelengths of 296 nm and 330 nm, respectively. Peaks were identified on the basis of the retention times of the standards α-, β-, γ- and δ-tocopherol separately and their concentrations were calculated using respective external calibration curves [23 (link)].
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